Saturday 11 April 2015

PUPULA DUPLEX or EVIL EYE

The phrase “pupula duplex” can be traced back to Ovid, and other ancient writers who referred to it as the “Evil Eye.”Although the term “pupula duplex” is practically non-existent in modern medical literature, a condition known as polycoria could roughly resemble two irises in certain cases. According to the Handbook of Pediatric Neuro-Ophthalmology (2006), polycoria is described as “a condition in which there are many openings in the iris that result from local hypoplasia of the iris stroma and pigment epithelium. True polycoria actually is a



condition in which there is more than one pupil and the multiple pupils all have a sphincter and the ability to contract.”t has been pointed out that “most cases of polycoria, however, are actually pseudopolycoria as only one of the pupils is the true pupil with an iris sphincter muscle. Therefore, in almost all clinical situations, the correct term is pseudopolycoria.”There are also other eye disorders or eye injuries which can give the illusion of multiple pupils.Since no official medical literature exists on this condition there is limited information available related to its possible causes. It is most likely a result of a genetic defect or mutation. It is not thought to be something that can be transmitted genetically. It isn’t contagious nor is it thought to be harmful to the person’s general health.

Wednesday 8 April 2015

ELISA- Enzyme Linked Immuno Sorbant Assay

In an ELISA, an antigen must be immobilized to a solid surface and then complexed with an antibody that is linked to an enzyme. Detection is accomplished by assessing the conjugated enzyme activity via incubation with a substrate to produce a measureable product. The most crucial element of the detection strategy is a highly specific antibody-antigen interactionELISAs are typically performed in 96-well (or 384-well) polystyrene plates, which will passively bind antibodies and proteins.The most commonly used enzyme labels are horseradish peroxidase (HRP) and alkaline phosphatase (AP). Other enzymes have been used as well, but they have not gained widespread acceptance because of limited substrate options. These include β-galactosidase, acetylcholinesterase and catalase. A large selection of substrates is available for performing the ELISA with an HRP or AP conjugate. The choice of substrate depends upon the required assay sensitivity and the instrumentation available for signal-detection (spectrophotometer, fluorometer or luminometer).


TYPES



Direct ELISA, Simple and Time-Saving

Initially in a direct ELISA test which is considered to be the simplest type of ELISA the antigen is adsorbed to a plastic plate, then an excess of another protein (normally bovine serum albumin) is added to block all the other binding sites. While an enzyme is linked to an antibody in a separate reaction, the enzyme-antibody complex is applied to adsorb to the antigen. After excess enzyme-antibody complex is washed off, enzyme-antibody bound to antigen is left. By adding in the enzyme's substrate, the enzyme is detected illustrating the siDirect ELISA, when compared to other forms of ELISA testing, is performed faster because only one antibody is being used and fewer steps are required. This can be used to test specific antibody-to-antigen reactions, and helps to eliminate cross-reactivity between other antibodies.
gnal of the antigen.

Disadvantages of direct ELISA

The primary antibody must be labeled individually, which can be time-consuming and inflexible when performing multiple experiments. Also, the signal is less amplified in direct ELISA, which means a lower sensitivity and could be viewed as a disadvantage to some


Indirect ELISA, conventional but efficient



Indirect ELISA is a two-step ELISA which involves two binding process of primary antibody and labeled secondary antibody. The primary antibody is incubated with the antigen followed by the incubation with the secondary antibody. However, this may lead to nonspecific signals because of cross-reaction that the secondary antibody may bring about.
1. Micro-well plates are incubated with antigens, washed up and blocked with BSA.
2. Samples with antibodies are added and washed.
3. Enzyme linked secondary antibody are added and washed.
4. A substrate is added, and enzymes on the antibody elicit a chromogenic or fluorescent signal.

Indirect ELISA advantages


  • High sensitivity:  More than one labeled antibody is bound per antigen molecule;
  • Flexible: Different primary detection antibodies can be used with a single labeled secondary antibody;
  • Cost-saving: Fewer labeled antibodies are required.





Sandwich ELISA, Highly Sensitive

Sandwich ELISA is a less common variant of ELISA,  but is highly efficient in sample antigen detection. Moreover, many commercial ELISA pair sets are built on this sanwich ELISA.
The sandwich ELISA quantify antigens between two layers of antibodies (i.e. capture and detection antibody). The antigen to be measured must contain at least two antigenic epitope capable of binding to antibody, since at least two antibodies act in the sandwich. Either monoclonal or polyclonal antibodies can be used as the capture and detection antibodies in Sandwich ELISA systems. Monoclonal antibodies recognize a single epitope that allows fine detection and quantification of small differences in antigen. A polyclonal is often used as the capture antibody to pull down as much of the antigen as possible. The advantage of Sandwich ELISA is that the sample does not have to be purified before analysis, and the assay can be very sensitive (up to 2 to 5 times more sensitive than direct orindirect ELISA), but lower than ELISpot.
Sandwich ELISA procedures can be difficult to optimize and tested match pair antibodies should be used. This ensures the antibodies are detecting different epitopes on the target protein so they do not interfere with the other antibody binding.
The steps are as follows:
  1. Prepare a surface to which a known quantity of capture antibody is bound.
  2. Block any nonspecific binding sites on the surface.
  3. Apply the antigen-containing sample to the plate.
  4. Wash the plate, so that unbound antigen is removed.
  5. A specific antibody is added, and binds to antigen (hence the 'sandwich': the Ag is stuck between two antibodies);
  6. Apply enzyme-linked secondary antibodies as detection antibodies that also bind specifically to the antibody's Fc region (non-specific).
  7. Wash the plate, so that the unbound antibody-enzyme conjugates are removed.
  8. Apply a chemical that is converted by the enzyme into a color or fluorescent or electrochemical signal.
  9. Measure the absorbency or fluorescence or electrochemical signal (e.g., current) of the plate wells to determine the presence and quantity of antigen.
Sandwich ELISAThe image at the bottom  includes the use of a secondary antibody conjugated to an enzyme, though, in the technical sense, this is not necessary if the primary antibody is conjugated to an enzyme. However, use of a secondary-antibody conjugate avoids the expensive process of creating enzyme-linked antibodies for every antigen one might want to detect. By using an enzyme-linked antibody that binds the Fc region of other antibodies, this same enzyme-linked antibody can be used in a variety of situations. Without the first layer of "capture" antibody, any proteins in the sample (including serum proteins) may competitively adsorb to the plate surface, lowering the quantity of antigen immobilized. Use of the purified specific antibody to attach the antigen to the plastic eliminates a need to purify the antigen from complicated mixtures before the measurement, simplifying the assay, and increasing the specificity and the sensitivity of the assay.

Sandwich ELISA Schematic Procedure:

(1) Plate is coated with a capture antibody; (2) sample is added, and any antigen present binds to capture antibody; (3) detecting antibody is added, and binds to antigen; (4) enzyme-linked secondary antibody is added, and binds to detecting antibody; (5) substrate is added, and is converted by enzyme to detectable form.

Sandwich ELISA advantages:

  • High specificity, since two antibodies are used the antigen/analyte is specifically captured and detected
  • Suitable for complex samples, since the antigen does not require purification prior to measurement
  • Flexibility and sensitivity, since both direct and indirect detection methods can be used

Competitive ELISA: Basic Principles

The central event of competitive ELISA is a competitive binding process executed by original antigen (sample antigen) and add-in antigen. The procedures of competitive ELISA are different in some respects compared with Indirect ELISA, Sandwich ELISA and Direct ELISA. A simplized procedure list is as follow:
  1. Primary antibody (unlabeled) is incubated with sample antigen.
  2. Antibody-antigen complexes are then added to 96-well plates which are pre-coated with the same antigen.
  3. Unbound antibody is removed by washing the plate. (The more antigen in the sample, the less antibody will be able to bind to the antigen in the well, hence "competition.")
  4. The secondary antibody that is specific to the primary antibody and conjugated with an enzyme is added.
  5. A substrate is added, and remaining enzymes elicit a chromogenic or fluorescent signal.
For competitive ELISA, the higher the sample antigen concentratiDiagram of Competitive ELISA Stepson, the weaker the eventual signal. The major advantage of a competitive ELISA is the ability to use crude or impure samples and still selectively bind any antigen that may be present.
(Note that some competitive ELISA kits include enzyme-linked antigen rather than enzyme-linked antibody. The labeled antigen competes for primary antibody binding sites with your sample antigen (unlabeled). The more antigen in the sample the less labeled antigen is retained in the well and the weaker the signal).
It is common that the antigen is not first positioned in the well.

Competitive ELISA  advantages:

  • High specificity, since two antibodies are used the antigen/analyte is specifically captured and detected
  • Suitable for complex samples, since the antigen does not require purification prior to measurement
  • Flexibility and sensitivity, since both direct and indirect detection methods can be used


Monday 6 April 2015

                                           ANTIPSYCHOTICS



















Psychiatric Uses Of Antipsychotic Drugs 

1. Schizophrenia: Acute and Chronic Maintenance 
2. Psychotic Depression (With Antidepressants) 
3. Acute Mania (With Lithium)
4. Autism (For Control of Aggressive Behaviors) 
5. Gilles de la Tourette’s Syndrome – Chronic Tics 
6. Severe Agitation In Mentally Retarded  
7.Alzheimer’s Patients


Pharmacological Actions of Antipsychotics At CNS Receptors

 Dopamine: Antagonists at D2 or Partial Agonist at D2             (aripiprazole) 
 • Serotonin: Antagonists at 5-HT2 A
 • Histamine: Antagonists at H1 
 • Cholinergic: Antagonists at muscarinic M1-4 
 • Noradrenergic: Antagonists at α 1



Typical Antipsychotics Have Antagonist Actions That Are Greater for the Dopamine D2 Than the 5-HT2A Receptor



Phenothiazines & Derivatives                                          
                                                                                                               
Chlorpromazine                                                                       Fluphenazine 
Perphenazine

Butyrophenones     
Haloperidol 







Atypical Antipsychotics Have Antagonist Actions that are Greater for 5-HT2A than D2

Risperidone 
Olanzapine 
Quetiapine 
Clozapine 
Ziprasidone



Dopamine Hypothesis Of Schizophrenia :An Increase in Dopaminergic Activity in CNS 


1. All Antipsychotics are DA Receptor Antagonists

2. Therapeutic effects correlated with D2 affinity 

3. Dopamine Agonists (e.g., Amphetamines) Exacerbate Schizophrenic Symptoms at Low Doses 

4. Higher Doses of Amphetamines Induce Paranoid Psychotic Reactions in Normal Individuals 

5. Evidence of Changes in Dopamine Receptors in Schizophrenia is Still Controversial




Blockade of Dopamine D2 Receptors 


Emotion - Reduces expression of emotion 
Cognitive functions – Decreases cognitive processes in prefrontal cortex 
Motor functions – Produces akinesia and symptoms of Parkinsonism
Endocrine function – Produces increased release of prolactin






Sunday 5 April 2015

              SYSTEMIC LUPUS ERYTHEMATOSUS


Systemic lupus erythematosus (SLE) is a chronic inflammatory disease that has protean manifestations and follows a relapsing and remitting course. More than 90% of cases of SLE occur in women, frequently starting at childbearing age


SYMPTOMS






PATHOPHYSIOLOGY



DIAGNOSIS



The diagnosis of SLE is based on a combination of clinical findings and laboratory evidence. Familiarity with the diagnostic criteria helps clinicians to recognize SLE and to subclassify this complex disease based on the pattern of target-organ manifestations.
The presence of 4 of the 11 American College of Rheumatology (ACR) criteria yields a sensitivity of 85% and a specificity of 95% for SLE.
When the Systemic Lupus International Collaborating Clinics (SLICC) group revised and validated the ACR SLE classification criteria in 2012, they classified a person as having SLE in the presence of biopsy-proven lupus nephritis with ANA or anti-dsDNA antibodies or if 4 of the diagnostic criteria, including at least 1 clinical and 1 immunologic criterion, have been satisfied.



ACR mnemonic of SLE diagnostic criteria

The following are the ACR diagnostic criteria in SLE, presented in the "SOAP BRAIN MD" mnemonic:
  • Serositis
  • Oral ulcers
  • Arthritis
  • Photosensitivity
  • Blood disorders
  • Renal involvement
  • Antinuclear antibodies
  • Immunologic phenomena (eg, dsDNA; anti-Smith [Sm] antibodies)
  • Neurologic disorder
  • Malar rash
  • Discoid rash
                             Testing
The following are useful standard laboratory studies when SLE is suspected:
  • CBC with differential
  • Serum creatinine
  • Urinalysis with microscopy
  • ESR or CRP results
  • Complement levels
  • Liver function tests
  • Creatine kinase assay
  • Spot protein/spot creatinine ratio
  • Autoantibody tests
                          Imaging studies
The following imaging studies may be used to evaluate patients with suspected SLE:
  • Joint radiography
  • Chest radiography and chest CT scanning
  • Echocardiography
  • Brain MRI/ MRA
  • Cardiac MRI
                           Procedures
Procedures that may be performed in patients with suspected SLE include the following:
  • Arthrocentesis
  • Lumbar puncture
  • Renal biopsy
                         Management



Management of SLE often depends on the individual patient’s disease severity and disease manifestations,although hydroxychloroquine has a central role for long-term treatment in all SLE patients.

Pharmacotherapy
Medications used to treat SLE manifestations include the following:
  • Biologic DMARDs (disease-modifying antirheumatic drugs): Belimumab, rituximab, IV immune globulin
  • Nonbiologic DMARDS: Cyclophosphamide, methotrexate, azathioprine, mycophenolate, cyclosporine
  • Nonsteroidal anti-inflammatory drugs (NSAIDS; eg, ibuprofen, naproxen, diclofenac)
  • Corticosteroids (eg, methylprednisolone, prednisone)
  • Antimalarials (eg, hydroxychloroquine)